This invention relates to an anticoagulant combination of LACI and sulfated polysaccharides and, more particularly, to a combination of LACI and heparin or similar such anticoagulant sulfated polysaccharides which exerts a synergistic anticoagulant action in whole plasma.
Blood clotting can be activated via the intrinsic or the extrinsic pathways. The intrinsic pathway begins with the contact phase which involves the interaction of factor XII, kallikrein, high molecular weight kininogen, a foreign surface and factor XI. The product of this reaction, factor XI.sub.a, converts factor IX to factor IX.sub.a, which subsequently hydrolyzes factor X to factor X.sub.a in the presence of activated factor VIII, phospholipid, and calcium. Alternatively, the extrinsic pathway is initiated when plasma factor VII/VII.sub.a, binds to tissue factor (TF; thromboplastin) to form a complex which proteolytically activates factors IX and X. Once factor X.sub.a is formed, either via the intrinsic or the extrinsic pathway, it can bind factor V.sub.a, phospholipid, and calcium to form the prothrombinase complex which converts prothrombin to thrombin. Ultimately, thrombin causes the fibrin clot to form.
Heparin has been widely used as an anticoagulant in clinical conditions. The anticoagulant effect of heparin is to a large extent a direct consequence of its catalytic action on the inhibition of thrombin by antithrombin III, and to a lesser extent its catalytic action on the inhibition by antithrombin III of other coagulation proteases including factors XII.sub.a, XI.sub.a, IX.sub.a, X.sub.a and kallikrein (1-4). In the absence of heparin, antithrombin III does not inhibit factor VII.sub.a (5-8). In the presence of heparin, factor VII, was reported to be resistant to inhibition (6) or inhibited 50% by antithrombin III in 11 min (7), 75-90 min (8) or 6 hours (5). Thus the rate of factor VII, inhibition by antithrombin III is so slow that antithrombin III is unlikely a physiological regulator of the TF/factor VII pathway in the presence or the absence of heparin (9). In addition to the antithrombin III-dependent inhibition of proteases of the intrinsic pathway, heparin can also exert anticoagulant action by displacing factor X.sub.a and prothrombin from the prothrombinase complex in an antithrombin III-independent fashion (10, 11).
In the past few years evidence has accumulated that regulation of the extrinsic pathway may primarily involve a plasma-derived protein called lipoprotein-associated coagulation inhibitor (LACI) (12). This protein also has been referred to as extrinsic pathway inhibitor (EPI) (13), or tissue factor inhibitor (TFI) (14). The inhibitor is capable of complexing with factor X.sub.a directly, and inhibits TF activity by formation of an inert TF/factor VII.sub.a /factor X.sub.a /Ca.sup.2+ /inhibitor complex (12). Following the purification of apparently related inhibitor from Hep G2 hepatoma (14), the cDNA coding for the protein was subsequently cloned (15). Recently, expression of recombinant protein has generated large quantity of protein for in vitro and in vivo use.
The isolation of LACI from the conditioned media of Hep G2 cells, SK-Hep-1 cells, and Chang liver cells also is disclosed in co-pending application Ser. No. 07/77,366, filed Jul. 23, 1987, and the cloning of the cDNA coding for the LACI protein also is disclosed in co-pending application Ser. No. 07/123,753, filed Nov. 23, 1987, now allowed.
References cited herein by numbers in parentheses are listed hereinbelow.